Application of next generation sequencing in clinical microbiology and infection prevention

Abstract

Current molecular diagnostics of human pathogens provide limited information that is often not sufficient for outbreak and transmission investigation. Next generation sequencing (NGS) determines the DNA sequence of a complete bacterial genome in a single sequence run, and from these data, information on resistance and virulence, as well as information for typing is obtained, useful for outbreak investigation. The obtained genome data can be further used for the development of an outbreak-specific screening test. In this review, a general introduction to NGS is presented, including the library preparation and the major characteristics of the most common NGS platforms, such as the MiSeq (Illumina) and the Ion PGM™ (ThermoFisher). An overview of the software used for NGS data analyses used at the medical microbiology diagnostic laboratory in the University Medical Center Groningen in The Netherlands is given. Furthermore, applications of NGS in the clinical setting are described, such as outbreak management, molecular case finding, characterization and surveillance of pathogens, rapid identification of bacteria using the 16S-23S rRNA region, taxonomy, metagenomics approaches on clinical samples, and the determination of the transmission of zoonotic micro-organisms from animals to humans. Finally, we share our vision on the use of NGS in personalised microbiology in the near future, pointing out specific requirements.

The challenges of designing a benchmark strategy for bioinformatics pipelines in the identification of antimicrobial resistance determinants using next generation sequencing technologies

“…The ECDC study reported the requirement for sufficient bioinformatics expertise as one of the important hurdles to a more general implementation of NGS for routine testing 2 . This observation has also been expressed in recent case studies and reviews 7– 11 .…”

Medical Management and Diagnostic Approaches

“…Other molecular methodologies such as nextgeneration sequencing, which allows for the simultaneous screening of hundreds of agents, will surely add to the arsenal of molecular techniques available to the diagnostician in the near future (Chiu, 2013;Deurenberg et al, 2017;Forbes et al, 2017;Kim et al, 2018;Simner et al, 2018). Molecular techniques are also applicable in many other areas of rat medicine and biology, including the detection of contaminants in tissue culture material and monitoring of genetic purity of inbred strains or genetically engineered rats (Bryda and Riley, 2008;Shimoyama et al, 2017).…”

Identification and Antibiotic‐Susceptibility Profiling of Infectious Bacterial Agents: A Review of Current and Future Trends

“…Although being a high‐resolution technique (single nucleotide), the associated high cost, complex workflow, need for quality control and interfering contamination events, render this technology still weak for daily use in clinical microbiology. However, several clinical microbiology laboratories already use it for rapid bacteria identification by 16S–23S rRNA sequencing, for tracking the source of infection outbreaks, for surveillance of pathogens or for other applications . Regrettably, in the case of plasmid‐mediated outbreaks, the direct repeats and insertions in the plasmids are often omitted from contigs, highlighting the need to apply alterations to the protocols .…”